ABSTRACT Friedreich's ataxia (FRDA; OMIM 229300) is an autosomal recessive neurodegenerative disease caused by reduced expression of the mitochondrial protein frataxin (FXN). Frequently, the level of FXN inversely correlates with severity of symptoms. The majority of FRDA patients are homozygous for large expansions of GAA repeat sequences in intron 1 of the FXN gene, while a fraction of patients are compound heterozygotes with a missense or nonsense mutation in one FXN allele and an expanded GAA repeat sequence in the other. Homozygous and compound heterozygous mutant genotypes both result in reduced levels of FXN mature protein when compared with heterozygous carriers and healthy controls. The most prevalent missense mutation within the FRDA patient population changes a glycine to valine at position 130 (G130V). Patients with the G130V mutation exhibit different clinical symptoms than patients with homozygous GAA expansions, including retained reflexes and a slower disease progression. Unexpectedly, we and others have demonstrated that the level of mature FXN protein is more prominently reduced in FRDA G130V patient samples compared to patient samples harboring homozygous expansions. Our preliminary data revealed that normal mitochondrial maturation processing of the FXN protein to its final form is perturbed by the G130V mutation, resulting in increased accumulation of an intermediate isoform. We hypothesize that the unprocessed FXN-G130V intermediate isoform is functional and compensates for the substantial reduction of mature FXN, thus alleviating the severity of symptoms and slowing disease progression in FRDA G130V patients. Currently, no models or reagents exist to distinguish FXN-WT and FXN-G130V proteins in FRDA G130V patient specimens, and it remains unknown if FXN-G130V is processed and/or functional in disease-relevant tissues. The goal of this proposal is to generate suitable models that discern endogenous FXN-WT and FXN-G130V proteins in FRDA G130V patient specimens and to assess the expression and function of the G130V protein under physiological conditions. To achieve this, we will pursue the following specific aims: 1.) Development of a FRDA FXN-G130V primary cell line model to enable studies that will determine the subcellular localization, processing, and function of the endogenous FXN-G130V protein. 2.) Development of mouse models to define effects of the G130V mutation on FXN protein expression and function in vivo. Realization of these aims will provide a mechanistic basis to specify the contribution of FXN-G130V isoforms to FRDA pathogenesis, which is necessary for the design of tailored therapeutic strategies.